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Multiplexed imaging with enzyme-mediated amplification

An imaging and counterstaining agent technology, applied in the preparation of test samples, microbial determination/inspection, instruments, etc.

Pending Publication Date: 2022-07-08
AKOYA BIOSCI INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] However, conventional immunohistochemistry (IHC) methods may not necessarily provide sufficient signal for detection, especially for immune targets that are weakly expressed or cannot be effectively targeted by existing IHC reagents

Method used

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  • Multiplexed imaging with enzyme-mediated amplification
  • Multiplexed imaging with enzyme-mediated amplification
  • Multiplexed imaging with enzyme-mediated amplification

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Experimental program
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Embodiment

[0207] To evaluate the above method, 5 micron thick tissue sections were cut from formalin-fixed, paraffin-embedded human tonsil blocks. Sections were deparaffinized, hydrated, and subjected to antigen retrieval with citrate buffer. The samples were then stained with CD20 (L20) antibody obtained from Akoya Biosciences, Inc. (Menlo Park, CA) following the staining instructions accompanying the antibody. The CD20 antibody used for staining was preconjugated to oligonucleotides (sequence BX015, obtained from Akoya Biosciences, Inc.) according to the manufacturer's instructions.

[0208] After staining, tissue sections were fixed in paraformaldehyde, ice-cold methanol and in immobilization reagent (obtained from Akoya Biosciences, Inc.), followed by washing. with 1x Tissue sections were equilibrated in 20% dimethyl sulfoxide (DMSO) in assay buffer (obtained from Akoya Biosciences, Inc.) for 10 minutes.

[0209] After equilibration, tissue sections were hybridized with 5 μL o...

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Abstract

A method for imaging an analyte in a sample comprises contacting a biological sample with a binding agent, where the binding agent comprises a binding moiety that binds to the analyte and a first nucleotide sequence, contacting the biological sample with a catalyst, where the catalyst comprises a second nucleotide sequence linked to an enzyme, and wherein the second nucleotide sequence hybridizes to the first nucleotide sequence, contacting the biological sample with a localization agent wherein the localization agent comprises a substrate complementary to the enzyme and a third nucleotide sequence linked to the substrate, and contacting the biological sample with a labeling agent wherein the labeling agent comprises a fourth nucleotide sequence linked to an optical label, wherein the fourth nucleotide sequence is hybridized with the third nucleotide sequence.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS [0002] This application claims priority to US Provisional Patent Application No. 62 / 908,540, filed September 30, 2019, the entire contents of which are incorporated herein by reference. Background technique [0003] In 1942, antibodies were first used in tissue section analysis to visualize pneumococcal antigens in biopsies of mouse organs infused with live bacteria. Since then, immunohistochemistry has become a mainstay in clinical diagnosis and basic research. [0004] However, conventional immunohistochemical (IHC) methods do not necessarily provide sufficient signal for detection, especially for immunological targets that are weakly expressed or cannot be effectively targeted by existing IHC reagents. [0005] SUMMARY OF THE INVENTION [0006] The present disclosure features methods, compositions of matter, and kits for immunohistochemistry of biological samples, wherein tyramide signal is amplified to enhance detection of...

Claims

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Application Information

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IPC IPC(8): C12Q1/6804
CPCC12Q1/6804C12Q1/6841G01N33/54306C12Q1/28C12Q2521/543C12Q2537/143C12Q2563/107C12Q2563/125C12Q2563/131C12Q2543/10C12Q2563/179C12Q2565/518C12Q1/6832G01N1/30
Inventor G.卡尔森
Owner AKOYA BIOSCI INC
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